immunocytochemical

Immunocytochemistry (ICC) is a laboratory technique that uses antibodies to detect specific antigens within individual cells, enabling visualization of the location and abundance of target molecules in a cellular context. The method is commonly applied to fixed, permeabilized cells prepared on slides, including cultured cells, cytospins, and smears.

In ICC, a primary antibody binds to the target antigen, and detection is achieved either directly with a labeled primary antibody or indirectly with a labeled secondary antibody. Detection methods include chromogenic visualization, where an enzyme such as horseradish peroxidase produces a colored precipitate, and fluorescent detection, which uses fluorophore-conjugated antibodies for fluorescence microscopy or confocal imaging. Indirect ICC generally provides higher sensitivity due to signal amplification.

Applications span basic research, clinical diagnostics, and quality control, including localization of proteins, assessment of cellular processes, and identification of cell types. ICC is related to immunohistochemistry (IHC) but is applied to single cells or cell preparations rather than tissue sections.

Key steps involve fixation to preserve structure, permeabilization to allow antibody access to intracellular epitopes, blocking to minimize non-specific binding, antibody incubation, and appropriate washing. Antigen accessibility can be influenced by fixation method and, for intracellular targets, may require antigen retrieval techniques.

Controls are essential and include negative controls (omitting the primary antibody), isotype controls, and positive controls to confirm antibody performance. Limitations include potential cross-reactivity and background staining, as well as dependence on epitope preservation by fixation. Multiplex ICC enables simultaneous detection of multiple targets, expanding its utility in profiling complex cellular phenotypes.